JP2003033173A - Dried microbial cell by spray-drying - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for producing a dried microbial cell or a composition containing the dried cell such as medicines or foods by which the death or injury of the microbial cells can be suppressed to maintained at a high survival rate of the microbial cells, and to provide a single micron-sized dried microbial cell having a large microbial cell number per unit weight and capable of being easily formed into the medicines or foods. SOLUTION: This method for producing the single micron-sized dried microbial cell comprises drying single micron-sized sprayed liquid drops with drying air. And the single micron-sized dried microbial cell is also provided.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for producing a dried microbial cell product using spray drying, a dried microbial cell product obtained by the production method, and a composition such as a medicine or food containing the dried microbial cell product. Regarding things.

[0002]

2. Description of the Related Art Freeze-drying or vacuum freeze-drying is often used as a drying method in the manufacturing process of medicines and foods. Freeze-drying is performed at about -20 ° C using liquid nitrogen or the like.
It is a method of drying at a low temperature of about 160 ° C., and vacuum freeze-drying is a method of drying under a pressure of about 35 ° C. or less and about 50 to 400 hPa.

However, when freeze-drying or vacuum freeze-drying is adopted as a drying method in the process of producing cells such as lactic acid bacteria and yeast, or a composition such as pharmaceuticals or foods containing the cells, the cells are killed during drying. There was also the drawback of being damaged. This is because it takes a long time to reach the freezing temperature in freeze-drying or vacuum freeze-drying, and the fungus is damaged by metabolites of the fungus or the freezing process generated during that time. In particular, vacuum freeze-drying has a very slow drying rate, and the above-mentioned problem becomes more prominent because the drying time becomes longer if all the water in the medicine or food is to be removed.

Further, in order to carry out freeze-drying or vacuum freeze-drying, a very large-scale equipment is required, and the equipment investment is also very expensive. Furthermore, if freeze-drying or vacuum freeze-drying is adopted as the drying method, it becomes difficult to manufacture in a continuous process, and the manufacturing process becomes complicated.
In addition, after freeze-drying or vacuum freeze-drying, further steps such as pulverization and homogenization are required, so that the number of manufacturing steps increases, and killing of bacteria during that time also poses a problem. Such problems have been pointed out for freeze-drying or vacuum freeze-drying.

On the other hand, as a drying method, spray drying is known in addition to freeze drying or vacuum freeze drying. Spray drying is a method in which a liquid is made into droplets by an atomizing device, and the droplets are brought into contact with a drying air of relatively high temperature to evaporate moisture and dry. However, conventional spray drying is generally unsuitable for the production of pharmaceuticals or foods containing viable bacteria, because the temperature of the drying air is high and the bacterial cells are killed by the heat of the drying air. Further, if the temperature of the drying air is lowered in order to solve this problem, there is a drawback in that a sufficiently dried microbial cell product cannot be obtained and both the yield and the stability decrease.

[0006]

The object of the present invention is to suppress the killing or damage of bacterial cells as much as possible in the process of producing a dried bacterial cell product or a composition such as a drug or food containing the same.
An object of the present invention is to provide a method for drying a bacterial cell liquid which can maintain a high viable cell rate. Another object of the present invention is to provide a single micron-sized dried microbial cell product which has a large number of microbial cells per unit weight and is easy to mold as a medicine or food. Still another object of the present invention is to provide a method for producing a bacterial cell composition which can greatly simplify the production process, and an apparatus according to the production method.

[0007]

Means for Solving the Problems The inventors of the present invention have made earnest studies to achieve the above object, and as a result, by drying single-micron spray droplets with a drying air, the problems in the conventional spray drying have been solved. I got the knowledge that I could solve it. That is, while the spray droplets formed in the conventional spray dryer are about 10 to 40 μm, when the spray droplets are very small, that is, single micron, the surface area per unit weight of the spray droplets is small. Since it becomes large, the contact with dry air is efficiently performed. Therefore, when it is dried with a drying wind having a temperature similar to that of conventional spray drying, the time required for drying the bacterial cell liquid is shortened, and the killing or damage of the bacterial cell can be suppressed as much as possible. Further, even if the temperature of the drying air is lower than the temperature of the drying air in the conventional spray drying, sufficient drying can be performed without lowering the yield or stability, and since the temperature of the drying air is low, The killing or damage of the cells due to heat can be suppressed as much as possible.

Further, the present inventors have found that the dried microbial cell product obtained by drying single-micron spray droplets with dry air has a small particle size and contains single-micron sized particles. I found out. Dried microbial cells with a small particle size have the advantage that they can be easily molded into pharmaceuticals or foods in dosage forms such as tablets, because the pressure applied to the cells during compression molding is less and static electricity is less likely to occur. .

That is, according to the present invention, (1) a dried product of single micron cells, and (2) a viable cell count of 1.0 × 10 ⁵ to
2.0 × 10 ¹² CFU / g, The dried bacterial cell product according to (1) above, (3) The viable cell count is 1.0 ×
10 ^{11 to} 2.0 × 10 ¹² CFU / g, dried bacterial cells, (4) bacterial cells are bifidobacteria, lactobacilli, lactococci, spore-forming lactic acid bacteria, saccharifying bacteria, yeast, Aspergillus, Actinomycetes, Bacillus toyoi, B. licheniformis, butyric acid bacterium, acetic acid bacterium or amino acid fermenting bacterium (1) to
The dried bacterial cell product according to (3), (5) the above (1) to (4)
The dried microbial cell according to (1), wherein the dried microbial cell is further coated or masked, and (6) the sprayed droplets of single micron are dried with dry air. (5) The method for producing a dried microbial cell, (7) The method for producing a dried microbial cell according to (6), wherein single-micron spray droplets are formed by a four-channel nozzle. Method, (8) The method for producing a dried microbial cell according to the above (6) or (7), characterized in that the drying air has a temperature of 5 to 150 ° C. at the inlet of the drying chamber.
(9) The fungus body according to the above (5), characterized in that a liquid coating agent or masking agent and a microbial cell liquid are simultaneously sprayed to form spray droplets, and the spray droplets are dried with a dry air. Method for producing dried product, (10) Coating of liquid by supplying bacterial cell liquid from one liquid flow channel in a four-flow channel nozzle having two liquid flow channels and two gas flow channels and from other liquid flow channels Agent or masking agent is supplied, compressed gas is supplied from two gas flow paths, and sprayed droplets generated are dried with a dry air. Method, (11) a step of performing fluidized bed granulation using a powdery substance and a binder, and a drying step of obtaining a dried product of single micron cells by spray drying are performed in the same container. Method for producing bacterial cell composition, (12) Fluidized bed construction using powder and binder And a drying step of obtaining a dried product of single micron cells by spray drying in the same container, (13) a granulated product comprising a powdery material and a binder , A bacterial cell composition characterized in that a dried product of single micron cells is attached, (14)
The dried bacterial cell product according to (1) to (5) or (13)
A medicine or food containing the bacterial cell composition according to (15), and (15) a single-micron droplet containing the bacterial cell,
Regarding

Further, according to the present invention, (1) single-micron spray droplets are dried with warm hot air to obtain a dried product of single-micron cells, and (2) single-micron spray droplets are formed by a four-passage nozzle. The dried microbial cell product according to (1) above,
(3) The dried bacterial cell product according to (1) or (2) above, wherein the temperature of the hot dry air in the drying chamber is 5 to 150 ° C.
(4) The dried or dried bacterial cell product according to (1) to (3), which is coated or masked, and (5) spray droplets are formed by simultaneously spraying a liquid coating agent or masking agent and a bacterial cell liquid. The dried bacterial cell product according to any one of (1) to (4) above, (6) a four-channel nozzle having two liquid channels and two gas channels,
Single micron spray droplets generated by supplying bacterial fluid from one liquid channel, supplying a liquid coating or masking agent from another liquid channel, and supplying compressed gas from two gas channels Was dried with warm dry air, and the dried bacterial cell product according to any one of (1) to (5) above, (7) the bacterial cell is a bifidobacteria, a lactobacillus, a lactococcus, a spore-forming lactic acid bacterium, or saccharification. Fungus, yeast, koji mold, actinomycete, Bacillus toyoi,
B. licheniformis, butyric acid bacterium, acetic acid bacterium, or amino acid fermenting bacterium, wherein the dried bacterial cell product according to any one of (1) to (6) above,
(8) The viable cell count is 1.0 × 10 ^{11 to} 2.0 × 10 ¹² C
FU / g, wherein the bacterium dried product according to the above (1) to (7), and (9) a medicament containing the microbial dried product according to the above (1) to (8). Also related to food.

[0011]

BEST MODE FOR CARRYING OUT THE INVENTION The dry cell product of the present invention is
Usually, it refers to individual dried cells or a collection of dried cells. The microbial cell dried product according to the present invention or the microbial cell dried product contained in the composition of the present invention such as a drug or food is any case as long as the microbial cell dried product having a particle size of single micron is included. Belongs to the present invention. Therefore, when the dried microbial cell is a mixture of a dried microbial cell having a particle size of single micron and a dried bacterial cell having a particle size of single micron or more, for example, when the average value of the particle size is single micron. Belongs to the present invention. Here, the single micron means a value of 1 to 10 μm by rounding off the first decimal place.

The cells used in the present invention are not particularly limited, but for example, Bifidobacterium bifidum, B. long
um, B. breve, B. adrecentis, B. infantis, B. pseud
Bifidobacteria such as olongum and B. thermophirum; for example, L
actobacillus acidophilus, L. casei, L. gasseri, L.
plantalum, L. delbrueckii subsp bulgaricus, L. del
Lactobacillus such as brueckii subsp lactis; eg Leucono
stoc mecenteroides, Streptococcus (Enterococcus) fa
ecalis, Streptococcus (Enterococcus) faecium, Stre
ptococcus (Enterococcus) hirae, Lactococcus lacti
s, Streptococcus thermophilus and other lactic acid bacteria; for example, Bacillus coagulans and other spore-forming lactic acid bacteria; for example, Bacillus subtilis, Bacillus mesentericus, Baci
Saccharomyces such as llus polyfermenticus; eg Saccaromy
yeasts such as ces cerevisiae, Candida; eg Aspergil
lus oryzae, Asp. nigar, Asp. Aspergillus oryzae such as Sojae; Strep
actinomycetes such as tomyces; for example, Bacillus toyoi, B. lic
Butyric acid bacteria such as heniformis and Clostridium butyricum; for example, acetic acid bacteria such as Acetobacter; amino acid fermenting bacteria such as Corynebacterim or other useful bacteria.

The above-mentioned bacterial cells can be obtained by culturing under known conditions or conditions similar thereto. For example, in the case of lactic acid bacteria, one or more of the lactic acid bacteria are usually cultured in a liquid medium containing glucose, yeast extract, peptone and the like at about 25 to 45 ° C. for about 4 to 24 hours, and then cultured. The bacterial cells are collected from the liquid and washed to obtain wet bacterial cells. The microbial cell may be a treated product of the microbial cell. Examples of the treatment include extraction with a suitable solvent. Specific examples include yeast extract, which is a hot water extract of yeast, and the like.

In the method for producing a dried microbial cell according to the present invention, first, the microbial cell is dissolved in a solvent to obtain a microbial cell liquid.
The solvent may be a known solvent used in the art, but water is preferable. If desired, ethyl alcohol and the like may be added. By adding ethyl alcohol etc.,
First, ethyl alcohol and the like are vaporized, and then water is vaporized, so that stepwise drying is possible. The cell fluid may be a suspension. As the solvent, a known solvent may be used as described above, but water or an aqueous solution obtained by adding ethyl alcohol or the like to water is preferable. When suspending, a suspending agent may be used. As the suspending agent, known ones such as sodium alginate or methyl cellulose may be used.

A protective agent may be added to the bacterial cell fluid.
The protective agent may be one commonly used in the art, for example, vitamins such as ascorbic acid; for example, amino acids such as glutamine, glutamic acid, L-cysteine, glycine, phenylalanine, serine or threonine; for example glucose, fructose. Saccharides or sugar alcohols such as sucrose, maltose, mannitol or maltitol; for example, oligosaccharides, polysaccharides such as cyclodextrin or dextrin; for example, fats such as higher fatty acids obtained from rapeseed, soybeans or peanuts; , Proteins obtained from milk or soybean, or proteolytic products such as peptides; inorganics such as magnesium sulfate; or others, such as sucrose fatty acid ester, malic acid, nucleic acids, yeast extract, skim milk powder,
Examples include peptone, gelatin, tannin and the like.
As the protective agent, the above substances may be used alone or in any combination of two or more kinds. Further, the protective agent is
It is preferable to add it in an amount of about 0.1 to 5.0 times, preferably about 0.5 to 3.0 times the wet cell weight.

Additives generally used in the art, such as an excipient, a binder, a disintegrant or an antistatic agent, may be further added to the above-mentioned bacterial cell fluid in a usual mixing ratio. As the excipient, for example, sugars such as lactose, granulated sugar or corn starch; sugar alcohols such as mannitol; cellulose such as natural cellulose or crystalline cellulose; or cellulose derivatives such as hydroxycellulose are used. Examples of crystalline cellulose include white powdery water-insoluble cellulose obtained by subjecting natural cellulose to acid treatment to subdivide the network molecular structure to some extent. Further, for example, a mixture of crystalline cellulose with carmellose, carmellose sodium or the like can be mentioned. Examples of the binder include hydroxypropylcellulose, hydroxypropylmethylcellulose, methylcellulose, dextrin, pregelatinized starch and the like. Examples of the disintegrant include cross-linked products such as carmellose calcium, carmellose or carmellose sodium (acdizol), cross-linked products such as polyvinylpyrrolidone (plasdone), corn starch or low-substituted hydroxypropyl cellulose. Examples of the antistatic agent include siloid, talc, aerosil and the like.

In the method for producing a dried bacterial cell product according to the present invention, the bacterial cell liquid is then subjected to spray drying. As the spray-drying device used for the spray-drying, a spray-drying device equipped with an atomizing device capable of forming single-micron spray droplets is preferable. With atomized droplets of very small particle size,
This is because the surface area per unit weight of the sprayed droplets becomes large, the contact with the dry air is efficiently performed, and the killing or damage of the bacteria due to the heat of the dry air can be suppressed as much as possible. Here, the single-micron spray droplets are those in which the particle diameter of the spray droplets is 1 to 10 μm rounded off to one decimal place.

As the spray drying device, specifically, for example, the atomizing device is a rotary atomizer (rotating disk), a pressure nozzle, or a two-pass nozzle or four-pass nozzle utilizing the force of compressed gas. A drying device may be used. In the present invention, any of the above spray drying devices may be used regardless of the type, but from the viewpoint that fine spray droplets can be obtained, it is preferable to use a spray drying device having a four-passage nozzle. preferable. The spray dryer may be one known per se.

As for the structure of the four-passage nozzle in the spray dryer having the four-passage nozzle, if the gas passage and the liquid passage are one system, two systems are symmetrically provided at the nozzle edge, and An inclined surface serving as a fluid flow surface is provided at the nozzle edge. One aspect of the structure of the four-passage nozzle will be described in more detail with reference to FIG. At the nozzle edge of the 4-channel nozzle, the liquid channel 3 or 4
The bacterial cell fluid that came out from the gas flow path 1 or 2
The thin liquid is thinly stretched on the fluid flow surface 5 by the high-velocity gas flow generated from the liquid, and the stretched liquid is atomized by the shock wave generated at the collision focus 6 at the tip of the nozzle edge to form a single-micron spray droplet 7. As described above, it is preferable that the four-passage nozzle adopts an external mixing method in which the compressed gas and the liquid are gathered at one point from both sides toward the collision focus at the tip of the nozzle edge. This is because this method enables long-term spraying without nozzle clogging.

As the compressed gas, for example, an inert gas such as air, carbon dioxide gas, nitrogen gas or argon gas can be used. In particular, when spray-drying an easily oxidizable substance, it is preferable to use an inert gas such as nitrogen gas or argon gas. The pressure of the compressed gas is about 1 to 15 kgf / cm ² , preferably about 3
It is about 8 kgf / cm ² . The amount of gas in the nozzle is about 1 to 100 L / min, preferably about 5 to 50 L / min, and more preferably about 10 to 20 L / min per 1 mm of the nozzle edge.

FIG. 2 shows one embodiment of a spray dryer having a four-passage nozzle. An embodiment of spray drying according to the present invention will be described with reference to FIG. 2. The bacterial cell fluid (reference numeral 21) is a four-channel nozzle (reference numeral 2) as described with reference to FIG.
8) Sprayed from. In the drying chamber (reference numeral 31), the sprayed droplets are brought into contact with the dry air sent from the dry air inlet (reference numeral 25) through the rectifier (reference numeral 33) to evaporate the water content and dry the microbial cell product. To get Temperature of the drying air at the entrance of the drying chamber (hereinafter referred to as "entrance temperature")
Is about 2 to 400 ° C, preferably about 5 to 250 ° C, more preferably about 5 to 150 ° C. Even if the inlet temperature is as high as about 200 to 400 ° C., the temperature of the material to be dried does not rise so much due to the heat of vaporization due to the evaporation of water, and the residence time in the drying chamber is shortened, so You can suppress death and damage as much as possible. The temperature of the drying air at the outlet of the drying chamber (hereinafter referred to as “outlet temperature”) is about 0 to 120 ° C., preferably about 5 to 90 ° C., and more preferably about 5 to 70 ° C. In the spray dryer shown in FIG. 2, the inlet temperature means the temperature measured by the inlet thermometer (reference numeral 26), and the outlet temperature is the temperature measured by the outlet thermometer (reference numeral 29). Say.

The drying chamber may be depressurized. By reducing the pressure, there is an advantage that the drying efficiency is improved and the drying time is shortened. The pressure of the drying chamber after depressurization varies depending on the size of the sprayed droplets and the like, and therefore cannot be generally stated. For example, the pressure in the low vacuum region, specifically, about 1.0 × 10 ³ to
It may be set to about 7.0 × 10 ³ Pa.

If a spray-drying apparatus having a four-passage nozzle is used, since the nozzle has two liquid passages as described above, two different bacterial cell fluids or bacterial cell fluids and other solutions or suspensions are used. By simultaneously spraying and from the different liquid flow paths, a dried microbial cell product in which these are mixed can be produced.
For example, by spraying the bacterial cell liquids of two different types of bacterial cells at the same time, a dried bacterial cell product containing the two types of bacterial cells can be obtained. Further, by spraying the bacterial cell liquid and the edible composition at the same time, a dried bacterial cell product containing the bacterial cell and the edible composition can be obtained.

Examples of the edible composition used in the present invention include liquid compositions of milk, meat, fish, fruits or vegetables. Alternatively, two or more of these may be mixed. These edible compositions are preferably concentrated to a water content of less than about 70% by weight prior to spray drying, leaving water to the extent that fluidity is not lost.

The above-mentioned liquid composition of fruits or vegetables includes, for example, seeds, roots, tubers, stems, leaves, derived from edible plants,
Examples include heated flowers or fruits or raw finely crushed parts. Preferred fruits or vegetables include, for example, Chinese chive,
Leaves of leeks, asparagus, fennel or cabbage; stems such as rhubarb or broccoli; seeds obtained from cocoa, peas, soybeans or cereals; roots such as carrots, onions, radish, celery or beets Tubers, such as cassava or potatoes; for example, tomato, kurujet, eggplant, banana, apple, apricot, melon, watermelon,
Fruits such as pears, plums, peaches, cherries, kiwi, sea buckthorn berry karin or mirabelle cherries;
Or a mushroom etc. are mentioned. Examples of the liquid composition of milk, meat or fish include milk, egg, meat or fish; heated or raw finely ground portions thereof; protein or protein hydrolyzate obtained therefrom.

Further, in the spray dryer having the four-passage nozzle, among the liquid passages having two passages in the nozzle,
By spraying the bacterial cell liquid from one liquid channel and spraying a liquid coating agent or masking agent from the other liquid channel, the bacterial cells can be coated or masked, and the coating or masking according to the present invention can be performed. The dried microbial cell product can be obtained. By coating the cells, the disintegration time can be adjusted, or the cells can be adjusted to act in a specific organ such as the intestine.
In addition, the appearance and quality can be improved. Further, in a drug or food containing two or more kinds of components, when the components are deteriorated due to contact with each other, the deterioration can be suppressed by coating one or both components. Moreover, bitterness and sourness can be suppressed by masking the bacterial cells. Particularly, a drug or food containing a high concentration of lactic acid bacteria or the like has a unique taste, and such a unique taste can be alleviated by coating or masking part or all of the surface of the bulk powder. There are advantages.

As the coating agent used in the present invention, for example, known materials having a coating ability such as hydroxypropyl cellulose (hereinafter abbreviated as HPC), starch paste, sugar solution such as silicone and candy, or various coating base materials. May be used. For example, enteric coated substrates include hydroxypropylmethylcellulose phthalate (HPMCP), hydroxypropylmethylcellulose acetate succinate (HPMCAS), cellulose acetate phthalate (CAP), carboxymethylethylcellulose (CMEC), methacrylic acid-
Examples thereof include acrylic acid ethyl ester copolymer and methacrylic acid-methacrylic acid methyl ester copolymer. Other coating substrates include ethyl cellulose, hydroxypropylmethyl cellulose, cellulose acetate phthalate, polyvinyl acetal diethylaminoacetate and the like. Examples of the masking agent used in the present invention include aminoacrylic methacrylate copolymer E, aminoacrylic methacrylate copolymer RS, and methacrylic acid copolymer L.
D or HPC etc. are mentioned. If these coating agents and masking agents are liquid, they can be used as they are. Alternatively, it may be diluted with a diluent such as water. If it is a solid, it is used after being dissolved or suspended in a solvent such as water.

Furthermore, instead of the coating agent or the masking agent, the microcapsules can be formed by spraying the wall film substance from one liquid channel of the four-channel nozzle. In this case, the microbial cell liquid is glycerin,
It is preferable to suspend the cells in a solvent inert to cells such as medium-chain triglyceride or corn oil. This is because if the solvent of the microbial cell liquid is water, the survival rate of the microbial cells may decrease due to the products produced by the active metabolism of the bacterium. As the wall membrane substance, for example, a cellulosic substance such as ethyl cellulose; for example, a polymer substance such as polylactic acid, polystyrene, polybutadiene, or styrene-methyl acrylate polymer; or, for example, a melting point above the body temperature of hardened palm oil or the like. The hardened oil etc. which it has are mentioned. If these wall film substances are liquid, they can be used as they are. Alternatively, it may be diluted with a diluent such as water. If it is solid,
The one dissolved or suspended in a solvent such as water is used.

As a method for producing the dried or coated bacterial cells or the method for producing microcapsules containing the dried bacterial cells according to the present invention, not only the above method but also the above-mentioned bacterial fluid and coating There is also a method in which the agent, the masking agent or the wall film substance is mixed in advance and the mixed solution is subjected to spray drying according to the present invention.

The dried bacterial cell product obtained by the above-mentioned spray drying according to the present invention is sent to the nozzle after considering whether the bacterial cell liquid is a solution or a suspension, or its properties such as concentration and viscosity. By adjusting the gas amount of the compressed gas and the liquid amount of the microbial cell liquid, a dried microbial cell product having an arbitrary particle size can be obtained. However, as described above, the particle size of the dried microbial cells is preferably about 1 to 10 μm by rounding off to the first decimal place. This is because, as the particle size is smaller, for example, static electricity is less likely to occur in the process of formulation, and less pressure is applied to the bacterial cells during tableting. The particle size of the dried microbial cells can be easily measured by a microscope. Further, in order to reduce the particle size of the obtained dried microbial cells, a treatment known per se may be performed before spray drying. Known treatments include treatments such as reducing the viscosity and surface tension of the microbial cell liquid, or reducing the primary particle size when the microbial cell liquid is a microbial cell suspension, or suspending it well. .

By reducing the particle size of the dried microbial cells as described above, there is an advantage that the viable cell ratio is increased and a drug or food having a large viable cell count can be provided. That is, it is preferable that the spray droplets are single micron in order to obtain a single micron dry cell product. If the particle size of the spray droplets is small, the surface area per unit weight of the spray droplets will be large, so that contact with the drying air will be performed efficiently, and even if the temperature of the drying air is relatively low, it will dry efficiently. Therefore, the killing or damage of the bacterial cells due to the heat of the dry air can be suppressed as much as possible. As a result, a viable cell ratio is increased and a dried cell product having a large viable cell count is obtained. Therefore, according to the spray drying according to the present invention, the viable cell ratio is about 1.0 × 10 ⁵ to about 2.
About 0 × 10 ¹² CFU / g, preferably about 1.0 × 1
^{0 11 ~2.0 × 10 12 CFU /} g , more preferably about from about ^{1.2 × 10 11 ~1.5 × 10 1} 2 CFU / g
Degree, most preferably about 1.5 × 10 ^{11 to} 8.0 × 1
A dried microbial cell product having a concentration of about 0 ¹¹ CFU / g can be obtained. Here, the measurement of the viable cell count in the dried bacterial cell differs depending on the bacterial cell, but can be easily measured by, for example, the method for quantifying each bacterial cell described in the Japanese Pharmacopoeia Standard for Pharmaceuticals.

In the present invention, the above spray drying and fluidized bed granulation may be combined. That is, using a device characterized in that the step of performing fluidized bed granulation using a powdery substance and a binder, and the drying step of obtaining a dried product of single micron cells by spray drying are performed in the same container. Then, granulation, spray drying of the bacterial cells, and mixing may be continuously carried out in the apparatus.
Conventionally, the steps of spray drying, granulation, and mixing have been performed using separate devices, but by performing granulation, spray drying, and mixing of bacterial cells in the same device, the manufacturing process is At the same time, simplification can be achieved, and at the same time, the mixing uniformity of the dried bacterial cell product and the granulated product is improved, and classification is less likely to occur.

As a specific embodiment of the apparatus according to the present invention, a binder spraying nozzle capable of spraying a binder from the upper part of the drying chamber to the lower part thereof, and a bacterial cell liquid from the upper part of the drying chamber to the lower part of the same. A fungus fluid spray nozzle that can be sprayed is installed, and a fluidized bed that can keep powdery materials in a fluidized state by blowing compressed gas or by sucking the air inside the apparatus with an exhaust fan There is a device attached to the bottom of the. Here, as the air forming the fluidized bed, air dried using a dehumidifying device known per se may be used. Here, in the apparatus, two spray nozzles are provided, and each spray nozzle is used as a binder spray nozzle or a bacterial fluid spray nozzle. Of course, only one spray nozzle is provided and the spray nozzles are combined. You may use together as a nozzle for spraying agent and a nozzle for spraying bacterial fluid.

Using the above apparatus, a step of performing fluidized bed granulation using a powdery substance and a binder and a drying step of obtaining a dried product of single micron cells by spray drying are performed in the same apparatus. Specific embodiments of the method for producing a bacterial cell composition characterized by the following are described below. In the following embodiments, the fluidized bed granulation step is performed first, and then the drying step is performed, but these steps may be performed simultaneously. First, fluidized bed granulation is performed using a powdery material and a binder. Specifically, powder is obtained by (a) blowing a compressed gas, preferably an inert gas such as nitrogen gas or carbon dioxide gas, from the bottom of the device, or (b) sucking the air in the device with an exhaust fan. The binder is sprayed from the binder spray nozzle while keeping the powder in a fluidized state, and the droplets of the binder are brought into contact with the fluidized powder to agglomerate the powder to granulate. To do. Here, as the nozzle for spraying the binder, a nozzle known per se may be used, but it is preferable to use the above-mentioned 4-channel nozzle. If a 4-channel nozzle is used, the spray droplets of the binder become smaller,
The particle size of the granulated product also becomes smaller. As a result, the time required for drying the flow-formed product is shortened, which leads to an increase in manufacturing efficiency. Further, by using the 4-channel nozzle, it is possible to obtain a granulated product having a relatively uniform particle size. As a result, the steps of crushing and sizing can be omitted, and classification is less likely to occur when the dried bacterial cell product and the granulated product are mixed. The particle size of the granulated product is
About 100 to 500 μm is preferable.

Next, the bacterial cell liquid is sprayed from the nozzle for spraying the bacterial cell liquid, and spray-dried to obtain a dried product of single micron bacterial cells. At this time, continuously from the previous stage, the compressed gas blown from the bottom of the device or the gas in the device sucked by the exhaust fan (hereinafter collectively referred to as "flowing gas") becomes dry air and the bacterial cell liquid At the same time as drying the sprayed droplets, the sprayed droplets are mixed with the granulated material in a fluidized state by the fluidizing gas. As an aspect of the obtained mixture,
Not only when the granulated particles and dried bacterial cell particles are independent and mixed, but also when the powdered particles and dried bacterial cell particles form one particle by the binder. ,
The case where the particles are mixed with the particles of the granulated product or / and the particles of the dried bacterial cell is included. In this way, it is possible to easily produce a bacterial cell composition characterized in that a dried product of single-micron cells adheres to a granulated product composed of a powdery substance and a binder. Here, as the nozzle for spraying the microbial fluid, a nozzle known per se may be used, but it is preferable to use the above-mentioned 4-channel nozzle. This is because if the four-passage nozzle is used, the advantages of the four-passage nozzle in the spray drying described above can be exhibited also in this manufacturing method.

One embodiment of the above apparatus and the method for producing the bacterial cell composition according to the present invention will be described with reference to FIG. For the fluidized bed granulation, the binder 4 is used in a state where the powdery material 48 is contained in the apparatus body 41 according to the present invention including the fluidized bed 42.
While spraying No. 5 from the binder spray nozzle 47, the exhaust fan is driven to continuously introduce the flowing gas 49 heated by the heat exchanger from the lower side of the apparatus main body 41. At this time, the flowing gas 49 may be dried in advance by using a dehumidifier (not shown). The flowing gas 49 introduced is sucked upward by the exhaust fan. The powder 48 is blown up from the fluidized bed 42 and suspended and suspended in the air as the wind speed of the fluidizing gas 49 increases, forming a so-called fluidized bed. Then, by bringing the droplets of the binder 45 into contact with the flowing powdery material 48, the powdery material 48 is aggregated, and granulation can be performed. Regarding spray drying, when the bacterial suspension 44 is sprayed using the bacterial fluid spray nozzle 46, the spray droplets of the bacterial fluid are dried by the fluidizing gas 49 that is pulled upward from the fluidized bed 42 by the exhaust fan. To be done. At this time, the flowing gas 4
If desired, lower the temperature of 9 from that of fluidized granulation to about 2
To about 400 ° C, preferably about 5 to 250 ° C, more preferably about 5 to 150 ° C, still more preferably about 2
The temperature is preferably about 0 to 70 ° C. This is to prevent the cells from being killed and damaged by the heat of the flowing gas as much as possible. The stream-formed product and the dried bacterial cell product produced as described above,
The flow gas 49 is mixed in the apparatus 41.

The powdery substance used for fluidized bed granulation is not particularly limited, but those which can be administered or ingested in combination with the dried bacterial cell product are preferred. Specific examples thereof include substances having a pharmacological action other than dried bacterial cells, excipients, and other finely powdered food materials. Those known per se can be used, or a mixture of two or more kinds may be used. Examples of the fine powdery food material include spices such as cinnamon, ginger, mustard, nutmeg, pepper, saffron, sesame, turmeric, parsley, mint, celery, onion, garlic, etc. Spices such as curry powder and mixed spices prepared by appropriately mixing spices and herbs; juice powders such as apple, orange, lemon, strawberry, pineapple, grapefruit, grape, melon, lime; coffee, green tea, black tea , Powdered beverages such as oolong tea, cocoa and barley tea; animal extract powders such as beef extract, pork extract, chicken extract, scallop extract, clam extract, oyster extract, crab extract, shrimp extract, bonito extract; garlic extract, onion extract, celery extract, cabbage Kiss,
Plant extract powder such as honey bean extract; powdered dairy products such as skim milk powder, whole milk powder, fermented milk powder; egg yolk powder, whole egg powder; cereal powder, animal and plant protein hydrolyzate, salt, amino acids, nucleic acid An example is a seasoning powder.

The binder used in the fluidized bed granulation is an additive that enhances the binding force between the particles of the powdery material or between the particles of the powdery material and the dried product of the bacterial cells, and a typical example is a water-soluble polymer. Can be mentioned. Specifically, for example, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, methyl cellulose and the like can be mentioned. In addition, as a binder, polyvinylpyrrolidone, gelatin,
It is also possible to use polyvinyl alcohol, starch paste or sugar solution. If desired, a compounding agent may be added to the binder. As the compounding agent, those known per se can be used, and for example, natural or synthetic surfactants such as lecithin, quillaja saponin, sucrose fatty acid ester, glycerin fatty acid ester, polyglycerin fatty acid ester, propylene glycol fatty acid ester; or Examples thereof include gelatin, casein, water-soluble peptide, soybean protein, albumin, and protein such as gluten.

The medicine or food containing the dried microbial cell according to the present invention may contain other components as long as it contains the dried microbial cell according to the present invention. For example, it may contain other pharmacologically active ingredients or other foods,
If desired, additives or additives having other desired functions may be included.

The dried microbial cell product obtained by spray-drying as described above is used as it is in the form of powder or granules, or in the form of tablets, pills or troches, and the dried microbial cell product of the present invention. It can be a medicine or food containing. When the drug or food containing the dried microbial cell of the present invention is in the form of powder or granule, the dried microbial cell obtained by the spray drying may be used as it is, and is used in the art. Processing known per se may be performed.

When the medicine or food containing the dried microbial cell according to the present invention is in the form of tablets, the dried microbial cell obtained by the spray drying is tableted by a tableting machine.
A known tableting machine such as a rotary tableting machine can be used. To the dried microbial cells obtained by spray drying according to the present invention, additives generally used in the art, such as a powdery or granular excipient, a binder, a disintegrant or an antistatic agent, are usually added. The mixture may be mixed in the compounding ratio of, and the resulting mixture may be tableted by a tableting machine. Further, the obtained tablets may be coated or masked with sugar coating, enteric coating or film coating. As the coating agent or masking agent, known agents such as those mentioned above may be used. The coating or masking can be performed by a known method or a method similar thereto. The tablet may be a sustained release tablet. As a sustained-release tablet,
Dried bacterial cells obtained by the spray drying, Gradumet dispersed in a plastic lattice that is discharged to the outside of the body, a wax in which the dried bacterial cells obtained by the spray drying are dispersed in a sponge-like wax lattice Examples thereof include a matrix, a resinate obtained by adsorbing a dried microbial cell product obtained by spray drying on an ion exchange resin, or a span tub composed of a plurality of layers having different dissolution and release properties. These sustained-release tablets can be manufactured by a known method or a method similar thereto.

When the drug or food containing the dried microbial cell of the present invention is in the form of a pill or troche, it can be produced in the same manner as a tablet. However, in the case of a lozenge, a disintegrant is not usually added because it is gradually dissolved or disintegrated in the mouth. Capsule may be prepared by filling capsules with the dried bacterial cells obtained by the spray drying. As the capsule base material, a known capsule base material such as gelatin may be used. In the case of soft capsules, a plasticizer such as glycerin or sorbitol may be used.
Further, the dried bacterial cells obtained by spray drying are compression-molded by a method such as tableting, which is optionally impregnated with fats and oils, and then coated with solid fats and oils, and then coated with a capsule base material. You may manufacture it. By doing this,
Even if water penetrates into the inside through the capsule coating, water does not directly touch the dried microbial cells, so that damage or death of the microbial cells due to water can be prevented. In addition, according to a conventionally known means, polylactic acid or the like may be used for microencapsulation.

The medicine or food containing the dried microbial cell of the present invention may be in a liquid dosage form such as a pharmaceutically acceptable dissolving agent, suspending agent, syrup or elixir. Examples of the diluent to be used include purified water, vegetable oil, ethanol and the like. In addition to the diluent, additives such as a wetting agent, a suspending agent, a sweetening agent, a flavoring agent, an aromatic agent or an antiseptic agent may be mixed.

The suspending agent is a well-known method in which the dried bacterial cell product obtained by spray drying is added to a solvent such as water or vegetable oil together with a suspending agent or other desired additives so that the whole becomes homogeneous. It is preferably produced by suspension by the method. Known suspending agents may be used, and examples thereof include gum arabic, carmellose sodium, methyl cellulose, sodium alginate and the like. The syrup is preferably produced by dissolving or suspending the microbial cell dried product obtained by the spray drying, together with sucrose and optionally other saccharides or sweeteners, in a diluent such as purified water.
A dry syrup may be used by dissolving or suspending the dried microbial cells at the time of use. The elixir is preferably produced by dissolving the dried bacterial cell product obtained by the above spray drying, if desired, together with an additive in ethanol having sweetness and aroma. The ethanol content is preferably about 4-40% by weight.

A medicine containing the dried microbial cell according to the present invention,
It may be prepared as a parenteral injection such as an aqueous injection, a non-aqueous injection, an aqueous suspension injection or a non-aqueous suspension injection. The injectable solution is obtained by dissolving the dried bacterial cells obtained by spray-drying in a solvent together with an additive, if desired, and then filtering or suspending in the solvent, then filling the container, sealing the container, and then sterilizing. It is preferable to carry out and manufacture. Examples of the solvent include aqueous solvents such as distilled water for injection, physiological saline and Ringer's solution; vegetable oils such as propylene glycol, polyethylene glycol or olive oil, and non-aqueous solvents such as alcohols such as ethanol. Examples of the additives include (a) stabilizers such as ethylenediamine, (b) preservatives such as paraoxybenzoic acid esters, benzyl alcohol, chlorobutanol, phenol or cresol, and (c) polyoxyethylene hydrogenated castor oil. A surfactant such as sucrose fatty acid ester, or a solubilizer such as lecithin or hydrogenated lecithin, (d) an isotonicity agent such as sodium chloride, glucose or glycerin, (e) a phosphate or citrate, for example. Buffering agents such as acid salts; or (f) suspending agents such as gum arabic, sodium carmellose, methylcellulose, or sodium alginate.

The medicine or food containing the dried microbial cell according to the present invention is used for promoting the health condition of mammals. It can also be used as an agricultural chemical or as an additive for feed. For example, lactic acid bacteria are widely known as one of the useful intestinal bacteria, and are considered to be deeply related to health. In particular, there are many reports on the physiological significance of bifidobacteria, lactococci, or lactobacilli, and the effect of inhibiting the growth of harmful bacteria by producing organic acids such as lactic acid or acetic acid in the intestine, vitamin production or immunity. It is clear that the activation of Therefore, many products such as pharmaceuticals and foods containing lactic acid bacteria have been developed for the purpose of maintaining and strengthening the health of humans, domestic animals, pet animals, etc. and preventing or treating diseases. Also,
Lactic acid bacteria are used because they are deeply involved in improving flavor and storability or maintaining quality in the production process of foods such as miso, soy sauce, pickles and sake.

The medicine containing lactic acid bacterium or bifidobacteria in the present invention is, for example, diarrhea, constipation, hypercholesterolemia, liver disease, immunosuppressant disease (immunosuppressant).
), Treatment or prophylaxis of enteritis (eg, gastroenteritis or colitis); intestinal after dysfunction due to food, drugs, chemicals or physical agents, or after surgical invasion, chemotherapy or contact infection Recovery; can be used to inhibit absorption of endotoxin and antagonize the production of endogenous toxins. This is because the drug acts to restore the intestinal mucosa and also regulate the intestinal enzyme potential when it changes as a result of stress conditions. In addition, it also has the functions of normalizing the function of the intestinal microflora, promoting synthesis of vitamins and proteins, promoting digestive or enzymatic processes and absorption processes, preventing colonization of pathogenic microorganisms, and stimulating immune response. is there.

The use of the dried lactic acid bacteria or bifidobacteria cells as a food product in the present invention is to obtain the dried bacterial cell product obtained by spray drying as described above, such as dairy products such as yogurt or cheese, and confectionery. And the like, and the like. Further, as the seasoning, the dried bacterial cell product obtained by the above spray drying can be used as it is.
Further, the dried microbial cells obtained by the spray drying may be dissolved or suspended in a solvent such as water or oil and used as a seasoning. Further, it may be mixed with other seasonings.

The lactic acid bacterium dried product obtained by the spray drying can also be used as a starter for stably producing a lactic acid fermented food on an industrial scale. The powdered lactic acid bacterium starter is used, for example, in dairy products, rye bread, fermented sausages, and the like.

The yeast microbial cell dried product obtained by spray drying can be used in the liquor industry such as beer, sake or wine by applying the property that yeast actively ferment alcohol. It can also be used for fermentation of bread and the like. Furthermore, yeast extract, which is a self-digesting solution or a hot water extract of yeast, contains a large amount of vitamins, organic bases or amino acids and has a high nutritional value. The dried product can be used as a medicine such as a vitamin preparation or a health food.

The dried bacterial cell product of Aspergillus oryzae obtained by the spray drying can be used for producing sake, amazake, shochu, miso, mirin, soy sauce and the like. Aspergillus oryzae is often used, but Asp.
igar is sometimes used. Further, aspergillus is often a strain that produces amylase, various enzymes such as protease and various organic bases, vitamins, etc. Therefore, the dried cell of Aspergillus oryzae in the present invention is used as a pharmaceutical or health food such as digestive enzyme preparation and vitamin preparation. Can be used.

[0052]

[Examples] [Example 1] [Culture of bacterial cells] A culture solution of Bifidobacterium bifidum pre-enriched with GAM medium (manufactured by Nissui Pharmaceutical Co., Ltd.) was inoculated into the medium at 0.1% by weight, and the mixture was incubated at 37 ° C. It was cultured for 16 hours. This was centrifuged, and the obtained precipitate was used as wet bacterial cells.

[Preparation of bacterial suspension] Mannitol is 0.5
M, sodium glutamate 0.5M, arginine 0.
The resulting wet bacterium was added to a dispersion medium prepared by adding dextrin to 35% by weight in an aqueous solution adjusted to a concentration of 1 M, sodium succinate 0.1 M, and magnesium sulfate 0.01 M. The body was suspended.

[Measurement of Number of Viable Bacteria in Bacterial Suspension] Measurement was carried out according to the quantification method of Bifidobacteria described in the section of the Japanese Pharmacopoeia Standard “Bifidobacteria”. That is, 1 mL of the bacterial suspension was taken, and diluted with a 10-fold dilution method using a diluent prepared as described below so that the number of viable bacteria in 1 mL was 20 to 200. Transfer 1 mL of this solution to a Petri dish,
To this, 20 mL of the agar culture medium for testing Bifidobacterium kept at 50 ° C. was added and rapidly mixed to solidify. This was anaerobically cultured at 37 ° C. for 60 hours, and the number of viable bacteria in the bacterial suspension was determined from the number of emerging colonies and the dilution ratio.

Here, in the agar medium for bifidobacteria test, of the following components, components other than equine blood were heated and melted, pH was adjusted to 7.2, and the contents were adjusted to 115 ° C. for 20 minutes using a high-pressure steam sterilizer. After heating and sterilization, it was cooled to 50 ° C. and horse blood was added to prepare. Bovine liver exudate 150 mL Animal meat peptone 10 g Casein peptone 5 g Yeast extract 5 g Meat extract 3 g Soybean peptone 3 g Glucose 10 g Potassium monohydrogen phosphate 1 g Potassium dihydrogen phosphate 1 g Starch 0.5 g L-Cysteine hydrochloride 0.5 g Magnesium sulfate 0 .2 g sodium chloride 0.01 g ferrous sulfate 0.01 g manganese sulfate 7 mg polysorbate 80 1 g agar 15 g horse blood 50 mL purified water 790 mL pH 7.1-7.3

The above diluted solution was prepared by mixing the following components and sterilizing by heating at 121 ° C. for 15 minutes using a high-pressure steam sterilizer. Anhydrous sodium monohydrogen phosphate 6.0 g Potassium dihydrogen phosphate 4.5 g Polysorbate 80 0.5 g L-Cysteine hydrochloride 0.5 g Agar 1.0 g Purified water 1000 mL pH 6.8-7.0

[Spray Drying] The bacterial suspension prepared as described above was spray dried using the spray dryer shown in FIG. 2 incorporating a 4-channel nozzle (described in Japanese Patent No. 2977080). That is, the bacterial suspension is fed from the two liquid flow paths to 8
Supply at a rate of mL / min (4 ml / min / 1 mm nozzle),
Compressed gas is supplied from two gas channels at a rate of 40 NL / min (value converted at 20 ° C.), and sprayed droplets are dried by a dry air to produce a dried microbial cell product according to the present invention. did. The drying air has an inlet temperature of 100 ° C.
It is supplied into the drying chamber at a rate of m ³ / min and the outlet temperature is 67 ° C.
Met.

[Measurement of Number of Viable Bacteria in Dried Bacteria] The number of viable bacteria in the obtained dried Bacteria was determined by quantification of Bifidobacterium described in the Japanese Pharmacopoeia Standard “Bifidobacteria”. It measured according to the method. That is, 5 g of the dried microbial cell product was taken in the same diluent as above, which had been preheated to 37 ° C., and the total amount was 50 mL. This was thoroughly stirred and then heated at 37 ° C. for 30 minutes to prepare a sample stock solution. This is diluted to 10
It was diluted by the double dilution method so that the number of viable bacteria in 1 mL was 20 to 200. 1 mL of this solution was placed in a Petri dish, and 20 volumes of agar culture medium for bifidobacteria test kept at 50 ° C. was placed therein.
mL was added and mixed quickly to solidify. This is 37 ℃
After anaerobically culturing for 60 hours, the number of viable bacteria in the dried bacterial cell product was determined from the number of emerging colonies and the dilution ratio.

The inlet temperature of the dry air is 80 ° C. and 60
The same procedure was performed by setting the temperature to 50 ° C.

Comparative Example 1 A bacterial suspension obtained in the same manner as in Example was freeze-dried. Freeze-drying condition is shelf temperature 30
The ultimate vacuum was 0.1 Torr (13.3 Pa) or lower. The viable cell count in the powder after drying was measured in the same manner as in Example 1.

Table 1 shows the viable cell counts and survival rates of Examples and Comparative Examples. Here, the survival rate was calculated from the following formula. Survival rate (%) = {(number of viable cells per 1 g of dried microbial cell x bacterial suspension solid content) / (number of viable cells per 1 g of bacterial suspension)} x
100

[Table 1] As shown in Table 1, it was found that by using the spray-drying according to the present invention, a dried microbial cell product having a higher survival rate than by using freeze-drying can be obtained.

[Particle size of dried microbial cell] In Example 1,
The dried bacterial cell product of the present invention obtained at an inlet temperature of 60 ° C. was used as a microscope (Digital H, manufactured by Keyence Corporation).
D microscope VH-7000). The particle size distribution was as follows.

[Table 2]

Example 2 In the same manner as in Example 1, bacterial cells were cultured to prepare a bacterial suspension, and the viable cell count in the bacterial suspension was measured. [Spray granulation] A fluidized bed granulator FLO-120 (manufactured by Freund Co.) was used, in addition to the conventional 2-channel spray nozzle, 4
A lactic acid bacterium-containing powder was prepared by spraying a lactic acid bacterium suspension into a fluidized bed using an apparatus incorporating a flow path nozzle (described in Japanese Patent No. 2977080). The process was roughly divided into two, and in the first process, a binder was sprayed on an excipient as a granular material using a two-passage nozzle to perform granulation.
In the second step, the lactic acid bacterium suspension was continuously spray-dried using a 4-channel nozzle while the fluidized bed was maintained at a low temperature, and at the same time, the lactic acid bacteria suspension was mixed with the granular material obtained in the first step. The operating conditions of the machine used in this experiment were a supply air volume of 70 m ³ / min and a nozzle air volume of 40 NL / min (value converted at 20 ° C.). In addition, the following compositions were used for the excipients and binders used for granulation.

The first step will be described in detail below. 120 kg of the above-mentioned shaping raw material is weighed and put into a hopper, the air supply temperature is set to 100 ° C., dry air is sent to form a fluidized bed, and a binder is used to liquid flow rate of 1.6 L /
Sprayed in minutes. At this time, the exhaust temperature was 45 to 50 ° C, and the temperature in the fluidized bed was 55 to 60 ° C. When the spraying of the binder was completed, it was dried in the fluidized bed as it was for 20 minutes to obtain a low moisture granulated product.

Next, the second step will be described in detail below. After the post-drying of the granulated product was completed, 350 mL of the bacterial suspension prepared in the same manner as in Example 1 was sprayed at a liquid flow rate of 16 mL / min using a 4-channel nozzle while the supply air temperature was lowered to 70 ° C. . At this time, the exhaust temperature was 30 to 35 ° C, and the temperature in the fluidized bed was 35 to 40 ° C. The lactic acid bacterium was pulverized by such spray drying at a low temperature, and simultaneously mixed in the fluidized bed to obtain the lactic acid bacterium-containing powder according to the present invention.

[Comparative Example 2] A lactic acid bacterium mixed powder was prepared so that the same amount of lactic acid bacterium was contained in the preparation. Example 1
2 g of the bacterial cell powder obtained in step 10 was added with corn starch.
Double the mixture to 20 g and add to 1180 g of the granules consisting of the excipient and binder obtained in the first step of Example 2,
A lactic acid bacterium mixed powder was obtained by mixing with a V-type mixer at 100 rpm for 3 minutes.

Powder 1 obtained in Example 2 or Comparative Example 2
The viable cell count contained in g was measured in accordance with the section of [Measurement of viable cell count in the dried bacterial cell product] of Example 1. In Example 2, the viable cell count was 5.12 × 10 ⁸ (CFU / g), and in Comparative Example, the viable cell count was 4.85 × 10 ⁸ (CFU / g). Therefore, by using the method according to the present invention in which spray drying and fluidized bed granulation are combined, a viable cell ratio of at least about the same as or more than the conventional method for producing a powder consisting of spray drying, granulation and a mixing step can be obtained. It was found that it is possible to simplify the manufacturing process while maintaining it.

[Example 3] The following components were added to 500 g of distilled water and stirred to prepare a coating agent. Composition of coating agent: Eudragit L30D-55 500 g Polyethylene glycol 6000 15 g Tween80 7.5 g Talc 45 g On the other hand, in the same manner as in Example 1, a bacterial suspension was prepared. Four
Spray drying was performed using the spray dryer shown in FIG. 2 in which a flow path nozzle (described in Japanese Patent No. 2977080) was incorporated.
That is, 4 having two liquid flow paths and two gas flow paths
Bacterial fluid is supplied from one liquid channel in the channel nozzle at a rate of 5 g / min, liquid coating agent is supplied from another liquid channel at a rate of 20 g / min, and compressed from two gas channels. The sprayed droplets produced by supplying gas at a rate of 40 NL / min were dried with a drying air to produce a partially coated microbial cell dried product according to the present invention. The inlet temperature of the dry air is 100 ° C., and the air is 0.9 mP at a rate of 1.0 m ³ / min.
It was supplied into the drying chamber of a. When 10 mg each of the conventional bulk powder of Bifidobacteria and the partially coated microbial cell dried product according to this example were taken and a taste test was conducted, the partially coated microbial cell dried product according to this example had an improved taste.

Example 4 100 g of the bacterial suspension prepared in the same manner as in Example 1 was mixed with an enteric coating base having the following composition, and the mixed solution was mixed with a 4-channel nozzle (Patent 2797080).
Spray drying was performed using a spray dryer shown in FIG. That is, the mixed liquid is supplied from the two liquid flow paths at a rate of 5.6 g / min, and the compressed droplets are supplied from the two gas flow paths at a rate of 40 NL / min. It was dried by air to produce the enteric-coated dried bacterial cell product of the present invention. The drying air had an inlet temperature of 80 ° C. and was supplied at a rate of 1.0 m ³ / min into a drying chamber of 0.9 mPa. Enteric coating base; AQOAT (manufactured by Shin-Etsu Chemical Co., Ltd.) 30 g Triethyl citrate 8.4 g Distilled water 161.6 g

[Acid resistance test of dried enteric coated bacterial cells] Acid resistance using the first liquid (pH 1.2, 36 to 38 ° C.) described in the Japanese Pharmacopoeia, General Test Method and Disintegration Test Method The test was conducted. The first liquid was placed in a container capable of keeping heat of about 1000 mL and kept at 36 to 38 ° C. Add the obtained dried microbial cells and stir well to add dried microbial cells 5, 1
After 5, 30, and 60 minutes, 1 mL was sampled. The sampled sample was diluted with the diluent described in Example 1 to be diluted to an appropriate concentration, and the viable cell count was measured in the same manner as in Example 1. In addition, the 1st solution of the pharmacopoeia is an artificial gastric juice, which is dissolved by adding 7 mL of hydrochloric acid and distilled water to 2.0 g of sodium chloride to make 1000 mL. In addition, the same test was performed on the conventional bulk powder of Bifidobacterium. FIG. 3 shows the relationship between the first liquid protection time and the viable cell count. In the case of conventional bulk powder, the viable cell count was 10 minutes after 5 minutes.
It was 1/00 or less, and no viable bacteria were observed after 30 minutes. On the other hand, about 1% of viable cells were observed in the coated bacterial bulk powder according to the present invention even 60 minutes after the addition. . That is, the acid resistance of the dried microbial cells was significantly improved by performing the above-mentioned coating.

[Example 5] Lactobacillus a as bacterial cells
A bacterial suspension was prepared in the same manner as in Example 1 except that cidophilus was used. The viable cell count in the bacterial suspension was measured in accordance with the method for quantifying lacumitone described in the section of the Japanese Pharmacopoeia Standard “Lacumitone”. That is, 1 m of bacterial suspension
Take 1 and use the diluted solution prepared as
It was diluted by the 0-fold dilution method so that the number of viable cells in 1 ml was 20 to 200. 1 ml of this solution was placed in a Petri dish, and 20 mL of the agar medium for lacumitone test kept at 50 ° C. was placed therein.
ml was added and mixed quickly to solidify. This is 37 ℃
After culturing for 48 hours, the number of viable bacteria in the bacterial suspension was determined from the number of appeared colonies and the dilution ratio.

Here, the agar medium for lacumitone test was prepared by mixing the following components and using a high-pressure steam sterilizer.
It was made by sterilizing by heating at ℃ for 15 minutes. Here, the tomato juice used was prepared by adding an equal amount of purified water to the tomato juice, boiling the mixture with occasional stirring, adjusting the pH to 6.8 and then filtering. Casein peptone 20 g Yeast extract 5 g Meat extract 15 g Glucose 20 g L-Cysteine hydrochloride 1 g Polysorbate 80 3 g Tomato juice 200 mL Agar 20 g Purified water 800 mL pH 6.8

The above diluted solution is mixed with the following components and sterilized by heating at 121 ° C. for 15 minutes using a high-pressure steam sterilizer.
Prepared. Sodium monohydrogen phosphate 6.0 g Potassium dihydrogen phosphate 4.5 g Polysorbate 80 0.5 g L-Cysteine hydrochloride 0.5 g Agar 1.0 g Purified water 1000 ml pH 6.9

The bacterial suspension prepared as described above was spray dried in the same manner as in Example 1. The operating conditions of the machine were Example 1 except that the inlet temperature was 60 ° C and the exhaust temperature was 45 ° C.
Was similar to. The number of viable bacteria in the obtained dried microbial cells was measured according to the method for quantifying lacumitone described in the section of the Japanese Pharmacopoeia Standard “Lacumitone”. That is, 5 g of the dried microbial cell product was taken up in a diluted solution preheated to 37 ° C. to make the total amount 50 mL. After stirring this well 37
The mixture was heated at 30 ° C for 30 minutes to prepare a sample stock solution. The number of viable bacteria in 1 mL is 20 to 20 by a 10-fold dilution method using a diluting solution.
Diluted to 0. 1 mL of this liquid was placed in a Petri dish, and 20 mL of the agar medium for the lacmitone test kept at 50 ° C. was added thereto and quickly mixed to solidify. This was anaerobically cultured at 37 ° C. for 48 hours, and the number of viable cells in the dried microbial cells was determined from the number of emerging colonies and the dilution ratio.

The number of viable bacteria in the dried microbial cells was 7.24 × 10.
¹¹ (CFU / g), and the survival rate was 90%.
The survival rate was determined in the same manner as in Example 1. Therefore, it was found that by using the spray drying according to the present invention, the survival rate of the microbial cells was increased, and as a result, a dried microbial cell product having an extremely large number of viable cells was obtained.

[0076]

EFFECT OF THE INVENTION By using a spray dryer equipped with an atomizer capable of forming single micron spray droplets, the surface area per unit weight of spray droplets increases,
Contact with dry air is efficiently performed. Therefore, when drying is performed with a drying air having a temperature similar to that of conventional spray drying, the time required to dry the bacterial cell liquid is shortened, and the killing or damage of the bacterial cells can be suppressed as much as possible. Further, even if the temperature of the drying air is lower than the temperature of the drying air in the conventional spray drying, sufficient drying can be performed without lowering the yield or stability, and since the temperature of the drying air is low, The killing or damage of the cells due to heat can be suppressed as much as possible. Therefore, by the method for producing a dried microbial cell according to the present invention, a dried microbial cell having a large number of viable cells can be obtained.

Further, the dried microbial cell product containing single micron-sized particles obtained by drying the sprayed droplets having a small particle size with a dry air as described above is a pressure applied to the microbial cell during compression molding or the like. It is easy to form into pharmaceuticals or foods in dosage forms such as tablets, since it requires less amount of electricity and static electricity is less likely to occur.

As a spray drying apparatus according to the present invention, by using a spray drying apparatus having a four-channel nozzle having two liquid channels and two gas channels, there are two liquid channels, so that a large amount of liquid can be obtained. In addition to being able to atomize and increase manufacturing efficiency,
By spraying two different solutions or suspensions simultaneously from different liquid flow paths, a mixed microbial cell dried product can be produced. Further, by spraying a liquid masking agent or coating agent from one liquid channel and spraying a bacterial cell liquid from the other liquid channel, a masked or coated bacterial cell dried product can be produced. Therefore, it is not necessary to separately carry out masking or coating, which reduces the number of manufacturing steps. Similarly, microcapsules can be produced by spraying the capsule base material at the same time as the bacterial cell liquid. Furthermore, by using a spray dryer having a four-passage nozzle, the particle size distribution falls within a relatively narrow range, so that subsequent steps such as sizing and sieving are easy to perform.

Spray drying is useful in terms of cost and resources because the scale of the apparatus is smaller than that of freeze-drying and the energy required for operating the apparatus is small. Further, the step of performing fluidized bed granulation using the powdery substance and the binder according to the present invention, and the step of obtaining a dried product of single micron cells by spray drying are performed in the same container. By continuously granulating, spray-drying, and mixing the cells in the device, the manufacturing process can be simplified, and at the same time, the mixing uniformity of dried cells and granules is improved. Classification is also less likely to occur.

[Brief description of drawings]

FIG. 1 shows an internal structure of a nozzle edge portion in a spray dryer having a four-passage nozzle.

FIG. 2 shows a schematic diagram of a spray dryer having a four-passage nozzle.

FIG. 3 shows the time-dependent changes in the viable cell count in the acid resistance test of the enteric-coated dried bacterial cell product.

FIG. 4 is characterized in that the step of performing fluidized bed granulation using the powdery substance and the binder according to the present invention and the step of obtaining a dried product of single micron cells by spray drying are performed in the same container. Fig. 2 shows a schematic diagram of the device.

[Explanation of symbols]

Gas flow path to which compressed gas is supplied Liquid flow path to which liquid containing the material to be dried is supplied 5 Fluid flow surface 6 collision focus 7 spray droplets 21 Bacterial fluid, coating agent or masking agent 22 Liquid flow path 23 gas flow path 24 Compressed air inlet 25 Dry air inlet 26 Entrance thermometer 27 atomizer 28 Spray nozzle 29 Outlet thermometer 30 Drying chamber pressure gauge 31 Drying room 32 Cell dry matter outlet 33 Rectifier 41 Device body 42 fluidized bed 43 Bug Filter 44 Bacterial fluid 45 binder 46 Nozzle for spraying bacterial fluid 47 Binder spray nozzle 48 powder 49 Flowing gas

─────────────────────────────────────────────────── ─── Continuation of front page (51) Int.Cl. ⁷ Identification code FI theme code (reference) A61P 1/14 A61P 1/14 3/02 3/02 (72) Inventor Koichi Hirayama Ibukidai, Nishi-ku, Kobe-shi, Hyogo Higashi-machi 7-chome 3-4 Biofermin Pharmaceutical Co., Ltd. Kobe Factory F-term (reference) 4B018 MD80 MD81 MD86 MD87 ME02 ME06 ME11 4B065 AA15X AA21X AA30X AA49X AA60X AA80X BD10 BD38 CA42 CA44 4C076 AA36 BB01 CC16 CC22 FF15 BC30C0 BC045A02 BC56 BC69 MA35 MA52 NA03 ZA66 ZC22

Claims (8)

[Claims] 1. A dried product of single micron cells. 2. The number of viable bacteria is 1.0 × 10 ^{5 to} 2.0 × 10.
^{It is 12} CFU / g, and the dried microbial cell product according to claim 1. 3. The cells are bifidobacteria, lactobacilli, lactococci, spore-forming lactic acid bacteria, saccharifying bacteria, yeasts, koji molds, actinomycetes,
Bacillus toyoi, B. The dried microbial cell product according to claim 1 or 2, which is licheniformis, butyric acid bacterium, acetic acid bacterium, or amino acid fermenting bacterium. 4. A dried microbial cell product, which is obtained by further coating or masking the dried microbial cell product according to claim 1. 5. A microbial cell composition characterized in that a dried product of single-micron microbial cells adheres to a granulated product comprising a powdery substance and a binder. 6. A medicine or food containing the dried microbial cell according to claim 1 or the microbial cell composition according to claim 5. 7. The medicine or food according to claim 6, which is a tablet. 8. A single-micron droplet containing cells.